Timothy P. White

Musculoskeletal adaptations to weightlessness and development of effective countermeasures

A Research Roundtable, organized by the American College of Sports Medicine with sponsorship from the National Aeronautics and Space Administration, met in November 1995 to define research strategies for effective exercise countermeasures to weightlessness. Exercise was considered both independently of, and in conjunction with, other therapeutic modalities (e.g., pharmacological nutritional, hormonal, and growth-related factors) that could prevent or minimize the structural and functional deficits involving skeletal muscle and bone in response to chronic exposure to weightlessness, as well as return to Earth baseline function if a degree of loss is inevitable. Musculoskeletal deficits and countermeasures are described with respect to: 1) muscle and connective tissue atrophy and localized bone loss, 2) reductions in motor performance, 3) potential proneness to injury of hard and soft tissues, and 4) probable interaction between muscle atrophy and cardiovascular alterations that contribute to the postural hypotension observed immediately upon return from space flight. In spite of a variety of countermeasure protocols utilized previously involving largely endurance types of exercise, there is presently no activity-specific countermeasure(s) that adequately prevent or reduce musculoskeletal deficiencies. It seems apparent that countermeasure exercises that have a greater resistance element, as compared to endurance activities, may prove beneficial to the musculoskeletal system. Many questions remain for scientific investigation to identify efficacious countermeasure protocols, which will be imperative with the emerging era of long-term space flight.

Satellite cell and growth factor involvement in skeletal muscle growth.

The activity of the satellite cell, discovered by Alexander Mauro, is of fundamental importance in postnatal skeletal muscle development, muscle adaptation to certain activity stimuli, and to muscle fiber regeneration following injury and transplantation operations. There are numerous mitogens and growth factors that influence satellite cell proliferation and differentiation in vitro and likely in vivo. The best understood purified growth factors are fibroblast growth factor (FGF), the insulin-like growth factors (IGF-I and -II), and transforming growth factor-beta (TGF-beta). Soluble extracts from injured muscle and chronically stretched muscle are also known to be mitogenic and are yet to be purified. Skeletal muscle development, hypertrophy, and regeneration can be viewed as points on a continuum with respect to the regulatory mechanisms of myogenic cell growth. The occurrence of fiber hyperplasia differs amongst some models of activity-induced growth and may reflect differences in the magnitude of the stimulus relative to the capacity of fibers to adapt. The relationships between the mechanical and environmental events coincident with an activity or injury stimulus and the role of specific muscle fiber satellite cell populations and growth factors are fertile areas for investigation. Insights from these experiments will yield a comprehensive understanding of the muscle growth process at the molecular, cellular, and tissue levels, and have implications for development and aging, health, disease, and adaptation.

Characteristics of Cat Skeletal Muscles Grafted with Intact Nerves or with Anastomosed Nerves

Grafting of 3-g extensor digitorum longus (EDL) muscles of cats may be made with (i) severence of the nerve with spontaneous reinnervation, termed standard grafts (ii) severence of the nerve with reinnervation facilitated by anastomosis of the nerve, termed nerve-anastomosed grafts; and (iii) preservation of the nerve, termed nerve-intact grafts. In previous studies, standard grafts developed a maximum isometric tetanic tension (P0) that was 22% of the value for control EDL muscles. We hypothesized that the low values of P0 resulted from incomplete reinnervation of muscle fibers. To test this hypothesis, EDL muscles were grafted in cats with nerves intact and with nerves anastomosed. In standard grafts differences were observed in both structure and function at 120 compared with 240 days after grafting. Characteristics of the nerve-intact and nerve-anastomosed grafts did not change significantly between 120 and 240 days and the data were pooled for comparisons with control EDL muscles. Nerve-anastomosed and nerve-intact grafts developed P0 values that were 34 and 64% of the control values, respectively. Nerve-intact grafts had a mass and fiber cross-sectional area not different from control EDL muscles. Compared with control values, all grafts had fewer fibers, more connective tissue, lower absolute and normalized P0, reduced capillary density, and increased fatigability. The greater P0 of nerve-intact compared with standard and nerve-anastomosed grafts supported our hypothesis that the degree of reinnervation is a factor that limits graft development. The presence of a necrotic core and the low tension development of even the nerve-intact grafts suggested that revascularization is a significant limitation as well.

Skeletal Muscle Regeneration and Plasticity of Grafts

The sequence of molecular and cellular events of muscle ontogeny leads to the proliferation, fusion, and differentiation of myoblasts to muscle cells. This sequence is closely paralleled in the grafting-ischemia model in which adult myoblast-satellite cells function as the muscle precursor cells. The study of skeletal muscle regeneration is a fertile and promising area of research in myogenesis. The early regenerative development and maturation of muscle is similar regardless of the perturbation that induced the degeneration-regeneration sequelae. In light of this, we maintain that the skeletal muscle graft model is useful to rigorously evaluate many regulatory aspects of skeletal muscle development and maturation in an adult animal host. One advantage of the graft model is that manipulation of the adult host, such as with exercise or hormone treatment, allows insight into their regulatory roles in muscle development and maturation. These approaches are often not possible for developing skeletal muscle in utero or in ovo. After skeletal muscle grafting, many structural and functional characteristics change with time until they reach a stable value. Successful regeneration requires revascularization, cellular infiltration, phagocytosis of necrotic muscle fibers, proliferation and fusion of muscle precursor cells, reinnervation, and recruitment and loading. The time taken to reach stable values varies among different structural and functional variables, and many reach stable values that are less than those of control skeletal muscle. There are differences in the degree of regenerative success because of the size of muscle mass grafted. In small and large grafts, regeneration is enhanced by facilitation of the reinnervation. Regeneration is evident without vascular repair in grafts of up to approximately 6 g, although in all but the 100 to 150-mg grafts in rats, a significant necrotic core is present. Regeneration is typically unsuccessful when muscle masses greater than 6 g are grafted without vascular repair. Large muscles can be grafted with vascular repair, and in this case, the cellular response is quite different, as the majority of fibers survive rather than degenerate and regenerate. Changing the components of physical activity during skeletal muscle regeneration can alter several attributes of the graft phenotype. The consensus of several experiments supports the interpretation that proper recruitment and force development by grafts are essential variables in the regulation of the development and maturation of muscle grafts. Morphological and physiological attributes of grafts adapt to changes in the habitual level of physical activity in a qualitatively similar fashion to control muscle.(ABSTRACT TRUNCATED AT 400 WORDS)

Myosin heavy chain phenotype in regenerating skeletal muscle is affected by thyroid hormone.

The purpose was to test hypotheses regarding the affect of thyroid hormone status on development of myosin heavy chain (MHC) protein phenotype in regenerating skeletal muscle. Soleus (SOL) and extensor digitorum longus (EDL) muscle grafts were studied 30 and 60 d following graft operations in eu-, hypo-, and hyperthyroid rats. Hyperthyroidism had a more profound affect on MHC phenotype than did hypothyroidism, and this was noted in SOL grafts more so than EDL grafts. For example, compared with euthyroid hosts at 30 d, SOL grafts from hyperthyroid hosts demonstrated a decrease from 83% to 3% in Type I MHC, and a decrease from 11% to 4% in Type IIa. Furthermore, Type IIx MHC increased from 5% to 12%, and Type IIb MHC increased from 3% to 82%. The change in Type I and IIb MHC noted at 30 d partially or fully reversed to euthyroid values by 60 d, whereas the hyperthyroid-induced differences in Type IIa and IIx were sustained. The adaptation of control muscles to hypo- and hyperthyroidism was qualitatively similar to muscle grafts for all MHC protein isoforms with the exception of Type IIx, which was expressed more so in grafts. For both grafts and control muscles, the adaptive response of MHC phenotype to sustained hyperthyroidism is transient for several of the MHC isoforms.

Skeletal Muscle Transplantation in Cats With and Without Nerve Repair

The regeneration of skeletal muscle fibers following free, whole muscle auto-transplantation has been well documented in rats (Carlson and Gutmann, 1975a und b). Neither vascular nor nerve repair was made, so revascularization and reinnervation occurred spontaneously or not at all. The time course of regeneration and the degree to which control values are restored have been described (Carlson et al., 1979). A limited number of studies have focused on the regeneration of skeletal muscle fibers following transplantation of skeletal muscles in larger species. Successful transplantations have been reported in cats (Hakelius et al., 1975; Faulkner et al., 1976; Maxwell et al., 1978; Faulkner et al., 1980) and monkeys (Markley et al., 1978; Markley and Faulkner, 1980; Maxwell et al., 1979). In dogs, Thompson (1971) has reported successful grafts whereas Lavine and Cochran (1976) and Watson and Muir (1976) described unsuccessful transplantations.

Alterations in skeletal muscle related to short-term impaired physical mobility

The purpose of this investigation was to describe and compare two methods of recovery of atrophied skeletal muscle following short-term impaired physical mobility. An animal model was used to study morphologic adaptations of atrophied soleus and plantaris muscles to the effects of 7 days of hind-limb suspension (HS) followed by either sedentary recovery or run training during a 28-day recovery period. Significant atrophy, demonstrated by decreased mean fiber area (MFA, in square micrometers), occurred during the 7-day period of HS. During recovery, MFA returned to control values 14 days earlier in the sedentary compared with the trained groups. Runs training following short-term atrophy induced by HS did not result in the high levels of frank muscle damage and type IIC fibers previously reported following long-term (28-day) atrophy.

Functional properties of palmaris longus muscles of rhesus monkeys transplanted as index finger flexors

This experiment with skeletal muscle autografts in monkeys was designed to retest previous findings that transplanted skeletal muscle can regenerate to a functional degree in primates without predenervation and to test a new hypothesis that increased functional demands on regenerated muscle grafts in monkeys may result in improved functional capacity of the grafts. Rhesus monkey index flexors were replaced with free palmaris longus muscle autografts with microneural anastomoses between the graft motor nerve and the severed profundus motor nerve. One monkey was taught selective index flexion before grafting and continued with this program after grafting to test the effect of training on the graft. Mature grafts were evaluated for in vivo contractile properties and by histology and histochemistry and were compared with a group of normal Rhesus palmaris longus muscles. The results reconfirm the capacity of nonpredenervated monkey skeletal muscle grafts to regenerate and to achieve some contractile ability and suggest that training of free muscle grafts may enhance recovery of their functional and structural properties.

Endurance-training induced changes in skeletal muscle phosphoglycerate kinase of old Wistar rats

Sufficiently intense, long-term, endurance training has been shown in several studies to induce a variety of adaptations in skeletal muscle, including a substantial restoration of the activities of several muscle enzymes which are known to be modified during biological aging. This activity-restoration may reflect either an increase in the amounts of enzyme proteins or an enhancement of the specific activities of these molecules. The present study examined the effect of long-term endurance training on the status of phosphoglycerate kinase in skeletal muscle of old rats, as compared with the enzyme isolated either from non-trained old or young animals. The kinetics of heat inactivation, which differ markedly between young and old forms of phosphoglycerate kinase, were used as a sensitive probe for the status of the enzyme. The results reveal a remarkable similarity between the heat inactivation patterns of phosphoglycerate kinase from the muscle of old, exercise-trained rats and enzyme purified from young animals, while enzyme samples isolated from sedentary old animals are significantly more heat-stable. Adaptation to endurance-training is thus evident at the molecular level, and maintains phosphoglycerate kinase in its young form. The aging of this enzyme has been previously shown to involve only conformational changes, which develop following a reversible partial oxidation of reactive cysteine residues. Whether the adaptation of the enzyme to endurance-training results from enhancement in its turnover rate (i.e., dwell time in the cell becoming too short for modifications to develop) or is due to increased protection against oxidation (being the first step in the enzymes aging) remains to be studied.

Mass and fiber cross-sectional area of soleus muscle grafts following training.

This study tested the hypothesis that endurance training initiated 28 d following grafting of the soleus muscle would increase fiber cross-sectional area concomitant with an increase in mass. Nerve-implant orthotopic grafting operations were performed on 6-wk-old rats anesthetized with pentobarbital sodium. A cohort of animals began running 28 d later. Control muscles were from age-matched, untrained rats. Mass and fiber cross-sectional area of grafts were 37 and 66% less than the respective control values at both 56 and 112 d post-grafting. Training increased graft mass by 49% over the non-run graft value of 82 +/- 8 mg at 56 d post-grafting. Continued training did not increase mass further. The grafts of trained rats were 33% greater than untrained at day 112 due to growth of grafts in untrained rats. Running had no effect on fiber cross-sectional area of grafts through 56 d, but by 112 d the cross-sectional area of Type I fibers was 30% greater than the non-run graft value of 1,271 +/- 81 micron2. By 112 d fiber type profiles were not different between control muscle and grafts from trained and untrained rats. We conclude that there is a dissociation between mass and fiber cross-sectional area in grafts compared to control muscle, and training affects these variables by similar magnitudes but at different times.